FR3095442A1 - GELIFYING LEGUMINOUS PROTEIN - Google Patents

Abstract

The invention relates to a legume protein composition having improved gel strength as well as its production method.

Description

GELLING LEGUM PROTEIN

The invention relates to the field of plant proteins, in particular legume protein isolates, even more particularly pea protein isolates.

Human daily protein requirements are between 12 and 20% of the food intake. These proteins are provided both by products of animal origin (meat, fish, eggs, dairy products) and by products of plant origin (cereals, legumes, seaweed).

However, in industrialized countries, protein intake is mainly in the form of protein of animal origin. However, numerous studies show that an excessive consumption of proteins of animal origin to the detriment of vegetable proteins is one of the causes of an increase in cancers and cardiovascular diseases.

Furthermore, animal proteins have many disadvantages, both in terms of their allergenicity, in particular concerning proteins derived from milk or eggs, and on the environmental level in relation to the misdeeds of intensive farming.

Thus, there is a growing demand from manufacturers for compounds of plant origin possessing advantageous nutritional and functional properties without however having the disadvantages of compounds of animal origin.

Soy was, and remains, the first plant-based alternative to animal protein. However, the use of soy has certain disadvantages. The soybean is more than frequently of GMO origin and obtaining its protein goes through a de-oiling step using solvent.

Since the 1970s, grain legumes, particularly peas, have developed strongly in Europe, mainly in France, as an alternative protein resource to animal proteins for animal and human food. The pea contains about 27% by weight of protein matter. The term “pea” is considered here in its broadest sense and includes in particular all wild varieties of “smooth pea” (“smooth pea”), and all mutant varieties of “smooth pea” and “wrinkled pea”. ("wrinkled pea"), regardless of the uses for which said varieties are generally intended (human food, animal nutrition and/or other uses). These seeds are non-GMO and do not require solvent deoiling.

Pea protein, mainly pea globulin, has been extracted and used industrially for many years. Mention may be made, as an example of a process for extracting pea protein, of patent EP1400537. In this process, the seed is ground in the absence of water (a process known as “dry grinding”) in order to obtain a flour. This flour will then be suspended in water to extract the protein.

Leguminous proteins, and in particular that of peas, are however much less gelling than those of soya. As presented in “Accessing gelling ability of vegetable proteins using rheological and fluorescence techniques” (Bastistaa & al., International Journal of Biological Macromolecules 36 (2005) 135–143, 2005), pea and lupine proteins are presented as less gelling than soy protein.

It is therefore of interest to obtain a legume protein, in particular a legume protein isolate, even more particularly a pea protein isolate whose gelling power or gel strength is improved.

General description of the invention

According to a first aspect of the invention, a legume protein composition is proposed, the legume being chosen in particular from peas, lupine and fava beans, characterized in that the gel strength of the protein composition according to test A is greater than 200 Pa, preferably greater than 250 Pa, even more preferably greater than 300 Pa and most preferably greater than 350 Pa. Preferably, the legume protein composition is a legume protein isolate and more preferably a pea protein isolate.

According to another aspect, there is proposed a method for producing a protein composition according to the invention, characterized in that it comprises the following steps:
1) implementation of legume seeds, preferably chosen between pea, lupine and horse bean;
2) grinding the seeds and producing an aqueous suspension;
3) separation by centrifugal force of the insoluble fractions;
4) coagulation of proteins by heating at isoelectric pH at a temperature between 55°C ⁺ / _- 2°C and 65°C ⁺ / _- 2°C, preferably 60°C ⁺ / _- 2°C, for a time between 3.5 min and 4.5 min, preferably 4 min;
5) recovery of the coagulated protein floc by centrifugation;
6) rectification of the pH to a value between 6 ^+/- _0.5 and 9 ^+/- _0.5 ;
7) optionally, heat treatment;
8) drying the coagulated protein floc;
9) grinding of the coagulated and dried protein floc in order to obtain a D90 particle size of less than 20 microns, preferably less than 15 microns, even more preferably less than 10 microns

According to a final aspect of the invention, the industrial uses, in particular animal and human food, of the legume protein composition, preferably of the legume protein isolate, chosen from pea, lupine and faba bean, are proposed. even more preferentially of the pea protein isolate according to the invention.

The invention will be better understood from the detailed description below.

Detailed description of the invention

According to a first aspect of the invention, there is therefore proposed a legume protein composition, the legume being in particular chosen from peas, lupine and fava beans, characterized in that the gel strength of the protein composition according to test A is greater than 200 Pa, preferably greater than 250 Pa, even more preferably greater than 300 Pa and most preferably greater than 350 Pa. Preferably, the legume protein composition is a legume protein isolate and more preferably a pea protein isolate.

The term "protein composition" should be understood in the present application as a composition obtained by extraction and refining, the said composition comprising proteins, macromolecules formed of one or more polypeptide chains consisting of the sequence of amino acid residues linked together by peptide bonds. In the particular context of pea proteins, the present invention relates more particularly to globulins (approximately 50-60% of pea proteins). Pea globulins are mainly subdivided into three subfamilies: legumes, vicilins and convicilins.

By "legume" will be understood in the present application the family of dicotyledonous plants of the order Fabales . It is one of the most important families of flowering plants, the third after Orchidaceae and Asteraceae by the number of species. It has about 765 genera comprising more than 19,500 species. Several legumes are important crops including soybeans, beans, peas, chickpeas, field beans, groundnuts, lentils, alfalfa, various clovers, broad beans, carob, licorice, the lupine.

By "gelling power" is meant the functional property consisting of the ability of a protein composition to form a gel or a network, increasing the viscosity and generating a state of matter intermediate between the liquid and solid states. The term "gel strength" can also be used. To quantify this gelling power, it is therefore necessary to generate this network and to evaluate its strength. To carry out this quantification, in the present invention, test A is used, the description of which is as follows:

1) Solubilization at 60° C. ^+/- ₂ ° _C. of the protein composition tested in water containing 15% ^+/- 2% in dry matter and at pH 7;
2) Stirring for 5 min at 60°C ^+/- ₂ °C;
3) Cooling to 20°C ^+/- ₂ °C and stirring for 24 hours at 350 rpm;
4) Implementation of the suspension in an imposed stress rheometer equipped with a concentric cylinder;
5) Measurement of elastic moduli G' and viscous modulus G'' by applying the following temperature profile:
To. _Phase 1: Measurement of parameter G'1 after stabilization at 20°C ^+/- ₂ °C and heating from a temperature of 20°C ^+/- ₂ °C to a temperature of 80°C ^+/- 2°C in 10 minutes;
b. Phase 2: stabilization at a temperature of 80°C ^+/- ₂ °C for 110 minutes; vs. Phase 3: cooling from a temperature of 80°C ^+/- ₂ °C to a temperature of 20°C ^+/- ₂ °C in 30 min and measurement of G'2 after stabilization at 20°C ^+/- ₂ °C;
6) Calculation of the gelling power equal to G'2 – G'1.

Preferably, the imposed stress rheometers are chosen from the DHR 2 (TA, instruments) and MCR 301 (Anton Paar) models, with a concentric cylinder-type rotor. They have a Peltier effect temperature control system. In order to avoid evaporation problems at high temperature, paraffin oil is added to the samples.

A "rheometer" within the meaning of the invention is a laboratory device capable of making measurements relating to the rheology of a fluid or a gel. It applies a force to the sample. Generally of small characteristic dimension (very low mechanical inertia of the rotor), it makes it possible to fundamentally study the mechanical properties of a liquid, a gel, a suspension, a paste, etc., in response to a force applied.

The so-called “imposed stress” models make it possible, by applying a sinusoidal stress (oscillation mode), to determine the intrinsic viscoelastic quantities of matter, which depend in particular on time (or on angular velocity ω) and temperature. In particular, this type of rheometer allows access to the complex modulus G*, itself allowing access to the moduli G' or elastic part and G'' or viscous part;

The first three steps consist of resuspending the protein in water, under specific conditions to maximize the subsequent measurement.

The water chosen is preferably reverse osmosis water, but drinking water can also be used.

Its temperature is 60°C ^+/- ₂ °C during the initial resuspension (1st and 2nd stages) then 20°C ^+/- ₂ °C after solubilization for 24 hours and cooling before measurement (3rd stage). In general and unless otherwise indicated, when a temperature is given in this description, it always includes a variation of ⁺ / _- 2°C, for example 20°C ⁺ / _- 2°C or 80°C ⁺ / _- 2°C.

A defined quantity of protein is added to said water in order to obtain a suspension titrating 15% ^+/- ₂ % in dry matter. To do this, equipment well known to those skilled in the art is used, such as beakers, magnetic bars. Shake a volume of 50mL for a minimum of 10 hours at 350 rpm at room temperature. In general, and unless otherwise indicated, the dry matter contents given in the present description always include a variation of ^+/- 2%, for _example 15% ^+/- ₂ %. The pH is adjusted to 7 ^+/- _0.5 using a pH meter and acid-base reagents, as is well known in the prior art.

The fourth step consists of introducing the sample into the rheometer, covering it with a thin layer of oil in order to limit evaporation.

The following temperature scale is then applied during the fifth stage: a. Phase 1: heating from a temperature of 20°C ^+/- ₂ °C to a temperature of 80°C ^+/- ₂ °C in 10 minutes; b. Phase 2: stabilization at a temperature of 80° C. ^+/- ₂ ° C. for 110 minutes; vs. Phase 3: cooling from a temperature of 80°C ^+/- ₂ °C to a temperature of 20°C ^+/- ₂ °C in 30 min.

The measurement of parameter G' is carried out continuously during this scale and is recorded.

The sixth and last step of test A consists in the exploitation of the recording. Two values are extracted: G'1 = value of G' at the start of phase 1 after stabilization at 20°C ^+/- ₂ °C and G'2 = value of G' at the end of phase 3 after stabilization at 20°C ⁺ / _- 2°C.

The gelling power is equal to G'2 - G'1.

Preferably, the legume protein composition according to the invention has a protein content greater than 80%, preferably greater than 85%, even more preferably greater than 90% by weight of dry matter relative to the total weight of the dry matter. .

The protein content is measured by any technique well known to those skilled in the art. Preferably, a dosage of the total nitrogen is carried out (in percentage by weight of nitrogen relative to the total dry weight of the composition) and the result is multiplied by the coefficient 6.25. This well-known methodology in the field of vegetable proteins is based on the observation that proteins contain on average 16% nitrogen. Any dry matter assay method well known to those skilled in the art can also be used.

Even more preferably, the protein composition has a D90 particle size of less than 20 microns, preferably less than 15 microns, even more preferably less than 10 microns.

By “D90”, is meant in the present invention the particle size in microns separating into two populations in number containing respectively 90% and 10% of all the total particles of the protein composition.

To perform this measurement of the D90, a laser particle sizer is preferably used, even more preferably the Mastersizer 2000 from the company Malvern. The parameters used are as follows: Use in liquid form, dispersion in ethyl alcohol; Refractive index: 1.52; Absorption index: 0.1; no use of ultrasound.

According to another aspect, a process is proposed which makes it possible to produce a legume protein composition according to the invention, characterized in that it comprises the following steps:
1) implementation of legume seeds, preferably chosen between pea, lupine and horse bean;
2) grinding the seeds and producing an aqueous suspension;
3) separation by centrifugal force of the insoluble fractions;
4) coagulation of proteins by heating at isoelectric pH at a temperature between 55°C ⁺ / _- 2°C and 65°C ⁺ / _- 2°C, preferably 60°C ⁺ / _- 2°C, for a time between 3.5 min and 4.5 min, preferably 4 min;
5) recovery of the coagulated protein floc by centrifugation;
6) rectification of the pH to a value between 6 and 9;
7) optionally, heat treatment;
8) drying the coagulated protein floc;
9) grinding of the coagulated protein floc in order to obtain a D90 particle size of less than 20 microns, preferably less than 15 microns, even more preferably less than 10 microns.

The method therefore starts with a step 1) of implementation of legume seeds, preferably chosen from pea, lupine and faba bean.

When the legume chosen is the pea, the peas used in step 1) will have been able to undergo beforehand steps well known to those skilled in the art, such as in particular cleaning (elimination of unwanted particles such as stones, dead insects, soil residues, etc.) or even the elimination of the outer fibers of the pea (outer cellulosic envelope) by a well-known step called “dehulling”.

Treatments aimed at improving the organoleptic such as dry heating (or roasting) or wet blanching are also possible. For bleaching, the temperature is preferably between 70°C ⁺ / _- 2°C and 90°C ⁺ / _- 2°C and the pH is adjusted between 8 ⁺ / _- 0.5 and 10 ⁺ / _- 0.5 , preferably at 9 ^+/- _0.5 . These conditions are maintained for 2 to 4 min, preferably for 3 min.

The method according to the invention comprises a step 2) of grinding the seeds and producing an aqueous suspension. If the grains are already in the presence of water, the water is conserved but can also be renewed, and the grains are ground directly. If the grains are dry, a flour is first made and this is suspended in water.

The grinding is carried out by any type of suitable technology known to those skilled in the art, such as ball mills, conical mills, helical mills, air jet mills or else rotor/rotor systems.

During the grinding, water can be added continuously or discontinuously, at the beginning, in the middle or at the end of the grinding, in order to obtain at the end of the stage an aqueous suspension of ground peas titrating between 15% and 25% by weight of dry matter (DM), preferably 20% by weight of DM, relative to the weight of said suspension.

At the end of grinding, a pH control can be carried out. Preferably, the pH of the aqueous suspension of ground peas at the end of _stage 2 _{is adjusted between 8 +/- 0.5 and 10 +/-} ^0.5 _, ^{preferentially} the pH is adjusted to 9 ^+/- 0.5 . The pH rectification can be carried out by adding acid and/or base, for example sodium hydroxide or hydrochloric acid.

The process according to the invention then consists of a step 3) of separation by centrifugal force of the insoluble fractions. These are mainly made up of starch and polysaccharides called "internal fibers". The soluble proteins in the supernatant are thus concentrated.

The method according to the invention comprises a step 4) of coagulation of the proteins by heating at isoelectric pH at a temperature between 55°C ⁺ / _- 2°C and 65°C ⁺ / _- 2°C, preferably 60°C ⁺ / _- 2 ° C, for a time between 3.5 min and 4.5 min, preferably 4 min. The aim here is to separate the pea proteins of interest from the other constituents of the supernatant of step 3). Such an example of a process is for example described in patent EP1400537 of the Applicant, from paragraph 127 to paragraph 143. are key in order to obtain a gelling protein composition according to the invention.

The following step 5) consists in the recovery of the coagulated protein floc by centrifugation. The solid fractions having concentrated the proteins are thus separated from the liquid fractions having concentrated the sugars and the salts.

In a step 6), the floc is resuspended in water and its pH is adjusted to a value between 6 ^+/- _0.5 and 9 ^+/- _0.5 . The dry matter is adjusted between 10% and 20%, preferably 15% by weight of dry matter relative to the weight of said suspension. The pH is adjusted using any acidic and basic reagent(s). The use of ascorbic acid, citric acid and potash, soda are preferred.

It is possible to carry out an optional step 7), consisting of a heat treatment aimed at guaranteeing the microbiological quality of the protein. This heat treatment can also be used to functionalize the protein composition. It is therefore preferably carried out according to a conventional scale of 100° C. ^+/- ₂ ° C. to 160° C. ^+/- ₂ ° C. for 0.01 s to 3 s, preferably between 1 and 2 seconds followed by immediate cooling.

In a step 8), the coagulated protein floc is dried to reach a dry matter greater than 80%, preferably greater than 90% by weight of dry matter relative to the weight of said dry matter. To do this, any technique well known to those skilled in the art, such as freeze-drying or even atomization, is used. Atomization is the preferred technology, especially multiple effect atomization.

The dry matter content is measured by any method well known to those skilled in the art. Preferably, the so-called “drying” method is used. It consists in determining the quantity of water evaporated by heating a known quantity of a sample of known mass: Weigh the sample at the start and measure a mass m1 in g; The water is evaporated by placing the sample in a heated enclosure until the mass of the sample stabilizes, the water being completely evaporated (preferably, the temperature is 105°C under atmospheric pressure), we weigh the final sample and a mass m2 in g is measured. The dry matter is obtained by the following calculation: (m2 / m1)*100.

The last step 9) is, like the previous step 4), key to obtaining the protein composition according to the invention. It consists of grinding the coagulated and dried protein floc in order to obtain a D90 particle size of less than 20 microns, preferably less than 15 microns, even more preferably less than 10 microns. Any type of technology known to those skilled in the art such as grinders with grinding wheels, balls, fine impacts or air jets will be potentially usable. However, the use of an opposed air jet mill is preferred, even more preferably the Netzsch CGS10. This type of shredder operates the reduction in size by generating collisions: the particles, accelerated by high-speed gas jets, are fragmented by impact.

According to a final aspect of the invention, the industrial uses, in particular animal and human food, of the legume protein composition, preferably of the legume protein isolate, chosen from pea, lupine and faba bean, are proposed. even more preferentially of the pea protein isolate according to the invention.

Due to its improved gelling power, the protein composition according to the invention is particularly suitable for food applications such as vegetable yoghurts or meat substitutes (“meat-analogs”).

The invention will be better understood using the non-limiting examples below.

Examples

Example 1: Production of a legume protein composition according to the invention

After decortication of the external fibers on a hammer mill, pea seeds are ground in order to obtain a flour. This is then soaked in water at the final concentration of 25% by weight of dry matter relative to the weight of said suspension, at a pH of 6.5, for 30 minutes at room temperature. The suspension of flour with 25% by weight of dry matter is then introduced into a battery of hydrocyclones, separating a light phase consisting of the mixture of proteins, internal fibers (pulps) and soluble and a heavy phase, containing the starch. The light phase leaving the hydrocyclones is then brought to a dry matter content of 10.7% relative to the weight of said suspension. The internal fibers are separated by passing through centrifugal decanters of the WESPHALIA type. The light phase at the centrifugal decanter outlet contains a mixture of proteins and solubles, while the heavy phase contains pea fibres.
The proteins are coagulated at their isoelectric point by adjusting the light phase at the outlet of the decanter centrifuge to a pH of 4.6 and heating this solution to 60° C. for 4 min. After coagulation of the proteins, a protein floc is recovered. This is resuspended at 15.1% dry matter relative to the weight of said suspension in drinking water. The pH of the suspension is rectified to a value of 7 with potassium hydroxide. Finally, a heat treatment is carried out at 130° C. for 0.4 s followed by flash cooling. The suspension is finally atomized on a NIRO multiple-effect MSD atomizer, the air inlet temperature being 180° C., and the outlet temperature being 80° C. The powder obtained derived 92.3% of dry matter relative to the total weight of the dry matter, including 85.5% of protein. This powder is called “Base for composition according to the invention”
This powder was then ground using a Netzsch CGS10 opposed air jet mill to obtain a powder with a D90 particle size of 7.3 microns.
The powdered protein composition obtained is called “Micronized protein composition according to the invention”.

Example 2: E x comparative example to demonstrate the influence of scale of heating of the protein composition during coagulation thereof

The purpose of this example is to demonstrate the impact of the coagulation scale on the functionalities of the protein composition according to the invention.

After decortication of the external fibers on a hammer mill, pea seeds are ground in order to obtain a flour. This is then soaked in water at the final concentration of 25.1% by weight of dry matter relative to the weight of said suspension, at a pH of 6.5, for 30 minutes at room temperature. The suspension of flour with 25% by weight of dry matter is then introduced into a battery of hydrocyclones, separating a light phase consisting of the mixture of proteins, internal fibers (pulps) and soluble and a heavy phase, containing the starch. The light phase leaving the hydrocyclones is then brought to a dry matter content of 11.2% relative to the weight of said suspension. The internal fibers are separated by passing through centrifugal decanters of the WESPHALIA type. The light phase at the centrifugal decanter outlet contains a mixture of proteins and solubles, while the heavy phase contains pea fibres.
The proteins are coagulated at their isoelectric point by adjusting the light phase at the outlet of the centrifuge decanter to a pH of 4.6 and heating this solution at 70° C. for 4 min. After coagulation of the proteins, a protein floc is recovered. This is resuspended at 14.9% dry matter relative to the weight of said suspension in drinking water. The pH of the suspension is rectified to a value of 7 with potassium hydroxide. Finally, a heat treatment is carried out at 130° C. for 0.4 s followed by flash cooling. The suspension is finally atomized on a NIRO multiple-effect MSD atomizer, the air inlet temperature being 180° C., and the outlet temperature being 80° C. The powder obtained derived 91.9% of dry matter relative to the total weight of the dry matter, including 84.9% of protein. This powder is called “Base for Comparative Protein Composition No. 1”.
This powder was then ground using a Netzsch CGS10 opposed air jet mill to obtain a powder with a D90 particle size of 8.2 microns.
The powdered protein composition obtained is called “Comparative Micronized Protein Composition No. 1”.

Example 3: Comparison of the different protein compositions obtained in Examples 1 and 2

In order to compare the protein composition, Test A is used as described in paragraph [0019] above, as well as the dry matter and the protein content:

Base for protein composition according to the invention Micronized protein composition according to the invention Basis for Comparative Protein Composition No. 1 Comparative Micronized Protein Composition #1 Dry matter (%) 92.3 96.7 91.9 96.5 Protein richness (% dry matter) 85.5 85.0 84.9 86.1 Gelling power (Pa) 98 375 103 109 D90 (in microns) 233.2 7.3 187.1 8.2

Table 1 above demonstrates unequivocally the extreme importance of the synergy of the coagulation temperature scale and the reduction of the particle size to a D90 particle size of less than 10 microns, in order to maximize the gelling power. The gelling power of the micronized protein composition according to the present invention is around 4 times higher than the base for protein composition according to the invention, the base for comparative protein composition No. 1 and the comparative micronized protein composition No. 1.

Claims (8)

Legume protein composition, the legume being chosen in particular from pea, lupine and horse bean, characterized in that the gel strength of the protein composition according to test A is greater than 200 Pa. Protein composition according to Claim 1, in which the legume protein composition is a legume protein isolate and more preferably a pea protein isolate. Protein composition according to Claim 2, characterized in that it has a protein content greater than 80%, preferably greater than 85%, even more preferably greater than 90% in dry matter relative to the total weight of the dry matter Protein composition according to Claims 1 to 3, characterized in that it has a D90 particle size of less than 20 microns, preferably less than 15 microns, even more preferably less than 10 microns. Process for the production of a protein composition according to claims 1 to 4, characterized in that it comprises the following steps:
1) implementation of legume seeds, preferably chosen between pea, lupine and horse bean;
2) grinding the seeds and producing an aqueous suspension;
3) separation by centrifugal force of the insoluble fractions;
4) protein coagulation heating at isoelectric pH at a temperature between 55°C ⁺ / _- 2°C and 65°C ⁺ / _- 2°C, preferably 60°C ⁺ / _- 2°C, for a time between 3.5 min and 4.5 min, preferably 4 min;
5) recovery of the coagulated protein floc by centrifugation;
6) rectification of the pH to a value between 6 ^+/- _0.5 and 9 ^+/- _0.5 ;
7) optionally, heat treatment;
8) drying the coagulated protein floc;
9) grinding of the coagulated and dried protein floc in order to obtain a D90 particle size of less than 20 microns, preferably less than 15 microns, even more preferably less than 10 microns. Process according to Claim 5, characterized in that the heat treatment consists of a scale of 100°C ^+/- ₂ °C to 160°C ^+/- ₂ °C for 0.01s to 3s, preferably between 1 and 2 seconds followed by immediate cooling. Process according to Claims 5 and 6, characterized in that the drying in step 8 is carried out by atomization, preferably by multiple-effect atomization. Process according to Claims 5 to 7, characterized in that the grinding in step 9 is carried out using a mill with opposed air jets.